The present invention relates to new types of terminally-activated polymeric materials which are useful in forming long-acting conjugates of bioactive materials. In particular, the invention relates to polymeric-based conjugates having increased therapeutic payloads and methods of preparing the same.
Over the years, several methods of administering biologically-effective materials to mammals have been proposed. Many medicinal agents are available as water-soluble salts and can be included in pharmaceutical formulations relatively easily. Problems arise when the desired medicinal agent is either insoluble in aqueous fluids or is rapidly degraded in vivo. Alkaloids are often especially difficult to solubilize.
One way to solubilize medicinal agents is to include them as part of a soluble product. Prodrugs include chemical derivatives of a biologically-active parent compound which, upon administration, eventually liberate the parent compound in vivo. Prodrugs allow the artisan to modify the onset and/or duration of action of an agent in vivo and can modify the transportation, distribution or solubility of a drug in the body. Furthermore, prodrug formulations often reduce the toxicity and/or otherwise overcome difficulties encountered when administering pharmaceutical preparations. Typical examples of prodrugs include organic phosphates or esters of alcohols or thioalcohols. See Remington""s Pharmaceutical Sciences, 16the Ed., A. Osol, Ed. (1980), the disclosure of which is incorporated by reference herein.
Prodrugs are often biologically inert or substantially inactive forms of the parent or active compound. The rate of release of the active drug, i.e. the rate of hydrolysis, is influenced by several factors but especially by the type of bond joining the parent drug to the modifier. Care must be taken to avoid preparing prodrugs which are eliminated through the kidney or reticular endothelial system, etc. before a sufficient amount of hydrolysis of the parent compound occurs.
Incorporating a polymer as part of a prodrug system has been suggested to increase the circulating life of a drug. However, it has been determined that when only one or two polymers of less than about 10,000 daltons each are conjugated to certain biologically active substances such as alkaloid compounds, the resulting conjugates are rapidly eliminated in vivo, especially if a somewhat hydrolysis-resistant linkage is used. In fact, such conjugates are so rapidly cleared from the body that even if a hydrolysis-prone ester linkage is used, not enough of the parent molecule is regenerated in vivo to be therapeutic.
Camptothecin and related biologically active analogs are often poorly water soluble and are examples of substances which would benefit from PEG prodrug technology. A brief overview of some previous work in the field is presented below.
Ohya, et al., J. Bioactive and Compatible PolymersVol. Jan. 10, 1995, 51-66, disclose doxorubicin-PEG conjugates which are prepared by linking the two substituents via various linkages including esters. The molecular weight of the PEG used, however, is only about 5,000 at most. Thus, the in vivo benefits are not fully realized because the conjugates are substantially excreted prior to sufficient linkage hydrolysis.
U.S. Pat. No. 4,943,579 discloses certain simple 20(S)-camptothecin amino acid esters in their salt forms as water soluble prodrugs. The reference does not, however, disclose using an amino acid as part of a linkage which would attach the alkaloid to a relatively high molecular weight polymer in order to form a prodrug. As evidenced by the data provided in Table 2 of the xe2x80x2579 patent, hydrolysis is rapid. Consequently, at physiologic pH, the insoluble base is rapidly generated after injection, binds to proteins and is quickly eliminated from the body before a therapeutic effect can be achieved. A related effort was directed to developing a water-soluble camptothecin sodium salt. Unfortunately, the water-soluble sodium salt of camptothecin remained too toxic for clinical application (Gottlieb et al,. 1970 Cancer Chemother, Rep. 54, 461; Moertel et al,. 1972 ibid, 56, 95; Gottlieb et al., 1972 ibid, 56, 103).
Commonly-assigned PCT publication WO 96/23794 describes bis-conjugates in which one equivalent of the hydroxyl-containing drug is attached to each terminal of the polymer. In spite of this advance, techniques which would further increase the payload of the polymer have been sought.
Thus, there continues to be a need to provide additional technologies for forming prodrugs of therapeutic moieties such as camptothecin and related analogs. The present invention addresses this need.
In one aspect of the invention, compounds of Formula (I) are provided: 
wherein:
J is 
E1-4 are independently selected from the group consisting of hydrogen, C1-6 alkyls, C3-12 branched alkyls, C3-8 cycloalkyls, C1-6 substituted alkyls, C3-8 substituted cycloalkyls, aryls, substituted aryls, aralkyls, C1-6 heteroalkyls, substituted C1-6 heteroalkyls, C1-6 alkoxy, phenoxy, C1-6 heteroalkoxy, 
and at least one of E1-4 includes a B moiety, wherein B is a leaving group, OH, a residue of a hydroxyl- or amine- containing moiety or 
wherein J1 is the same as J, or another member of the group defining J and E5 is the same as E1-4, or another member of the group defining E1-4;
Y1-2 are independently O or S;
M is a heteroatom selected from either X or Q; wherein X is an electron withdrawing group and Q is a moiety containing a free electron pair positioned three to six atoms from C(=Y2);
R2-5 and R7-8 are independently selected from the group consisting of hydrogen, C1-6 alkyls, C3-12 branched alkyls, C3-8 cycloalkyls, C1-6 substituted alkyls, C3-8 substituted cycloalkyls, aryls, substituted aryls, aralkyls, C1-6 heteroalkyls, substituted C1-6 heteroalkyls, C1-6 alkoxy, phenoxy and C1-6 heteroakoxy;
(m1) and (m2) are independently zero or one;
(n1), (n2), (p1), (p2) and (q) are independently zero or a positive integer;
Z is an electron withdrawing group; and
R1 is a polymeric residue such as a water-soluble polyalkylene oxide, preferably having a molecular weight of at least about 20,000 Daltons.
In preferred aspects of the invention, the polymeric residue is also substituted on the distal portion with another branching group to provide compounds of the formula (Ixe2x80x2): 
where all variables are as previously defined. The bifunctional compounds are thus formed when the polymeric residue (R1) includes both an alpha and an omega terminal linking group so that two, four or more equivalents of a biologically active agent, drug or protein, designated herein as B, can be delivered. Multifunctional compounds represented by the formula (Ixe2x80x2) are preferred.
When B is a residue of a hydroxyl-containing moiety or an amine-containing moiety, each B is attached via a hydrolyzable linkage which attaches to the polymer residue terminus.
Examples of hydroxyl-containing moieties for which on or more of improved aqueous solubility, decreased antigenicity, prodrug and/or controlled release delivery is desired include chemotherapeutic compound residues such as anti-fungal compounds, including triazoles, echinocandins, pneumocandins, etc, anti-cancer compounds such as camptothecin, paclitaxel, etoposide, anti-cancer platinum compounds containing OH groups, floxuridine or podophyllotoxin. In still further embodiments, other oncolytic agents, non-oncolytic agents such as anti-inflammatory agents, including steroidal compounds, as well as therapeutic low molecular weight peptides such as insulin are also contemplated.
Examples of amine-containing moieties for which one or more of improved aqueous solubility, decreased antigenicity, prodrug and/or controlled release delivery is desired include antimetabolites such as Ara-C or gemcitabine.
Alternatively, B can be a leaving group such as N-hydroxybenzotriazolyl, N-hydroxyphthalimidyl, halogen, xcfx81-nitrophenoxy, imidazolyl, N-hydroxysuccinimidyl, thiazolidyl thione, or other activating groups.
For purposes of the present invention, the term xe2x80x9cresiduexe2x80x9d shall be understood to mean that portion of a biologically active compound which remains after the biologically active compound has undergone a substitution reaction in which the prodrug carrier portion has been attached.
For purposes of the present invention, the term xe2x80x9calkylxe2x80x9d shall be understood to include straight, branched, substituted, e.g. halo-, alkoxy-, and nitro- C1-12 alkyls, C3-8 cycloalkyls or substituted cycloalkyls, etc.
For purposes of the present invention, the term xe2x80x9csubstitutedxe2x80x9d shall be understood to include adding or replacing one or more atoms contained within a functional group or compound with one or more different atoms.
The term xe2x80x9csufficient amountsxe2x80x9d for purposes of the present invention shall mean an amount which achieves a therapeutic effect as such effect is understood by those of ordinary skill in the art.
One of the chief advantages of the compounds of the present invention is that the prodrugs have a higher payload per unit of polymer than previous techniques. Another advantage is that the linkers achieve a proper balance between the rate of parent drug-polymer linkage hydrolysis and the rate of clearance of prodrug from the body. The linkages between the polymer and the parent compounds, also referred to herein as biologically-active nucleophiles, hydrolyze at a rate which allows a sufficient amount of the parent molecules to be released in vitro before clearance of the prodrug from the plasma or body. The high payload polymeric conjugates of the present invention are thus unique delivery systems which can contain up to four or a greater number of molecules of a drug.
Methods of making and using the compounds and conjugates described herein are also provided.